Project Summary Eosinophilic esophagitis (EoE) is a Th2 disease of increasing incidence and prevalence that results in clinical dysphagia due to esophageal rigidity and strictures. Esophageal stiffness occurs, in part, due to fibrosis causing a rigid extracellular matrix (ECM). Alzheimer's Disease (AD) is the most common cause of dementia in the US affecting 5 million Americans over the age of 65. In AD, ECM rigidity occurs due to amyloid which, together with neurofibrillary tangles, creates a neurotoxic ECM. One difficulty in studying AD is the paucity of human tissue and cells. In contrast, EoE requires repeated esophageal biopsy for diagnosis and maintenance, allowing for the repeated isolation and study of human tissue and cells. Our new proteomic analysis of EoE versus normal fibroblasts demonstrates a significantly dysregulated secretory and ECM network with 50 differentially expressed proteins (DEPs). Interestingly, 79% of the upregulated EoE fibroblast proteome represents genes that are also expressed in mature and/or fetal astrocytes (brainarnaseq.org), supporting the hypothesis that EoE and AD have parallel disease mechanisms. Further, an analysis of the protein interaction networks surrounding each of the 50 differentially expressed proteins in the EoE proteome and overlaid with an AD RNA sequencing data set showed that collagen triple helix repeat containing-1 (CTHRC1) had the strongest perturbed network and correlated positively with amyloid deposition (FDR adjusted p <0.001) and dementia Braak scores (FDR adjusted p<0.05). The machinery support scores for genes supporting the secretion of the EoE differentially upregulated protein, CD44, also correlated significantly with the expression of genes increased with amyloid deposition (p=0.01). The CD44 binding protein, hyaluronic acid (HA), is elevated in the AD brain as well as in allergic disorders such as asthma. Together, these findings led us to the central hypothesis that studies of the EoE fibroblast-ECM interactome and its specific components CTHRC1, HA, and CD44 will reveal pathogenic molecular mechanisms common to both EoE and AD. We further posit that this is due to parallels between brain astrocytes and esophageal fibroblasts. We will use primary esophageal cells and biopsies from patients with and without EoE to understand the CTHRC1 and CD44 interacting proteins using cutting edge in situ proteomic methodologies. We will validate the expression of the CTHRC1 and CD44/HA interactomes in biopsies of normal and EoE esophagus and normal and AD brains. We will evaluate the effects of the esophageal ECM on astrocyte function and understand the effects of HA and CTHRC1 on esophageal fibroblast and brain astrocyte function. Since EoE cells and biopsies are repeatedly procured, we aim to use EoE as a paradigm disease to decipher currently unappreciated disease mechanisms in AD. To successfully complete this proposal, we have assembled a collaborative team of investigators with expertise and experience in AD, translational EoE, proteomics, and systems biology.